Neural Tract-tracing Nucleic Acid Carriers Existing transfection reagents perform very poorly in mature neurons in vitro, and are not suitable for in vivo use. This Phase I project will test the performance of a novel nucleic acid carrier under development at Innovative Surface Technologies, Inc. (ISurTec) for universal transfection of neurons in vitro and in vivo. This technology is platform-based and consists of a polymeric nanoparticle for high-efficiency non-viral gene and siRNA transfection, chemically masked as a neural tract-tracer through surface modification. Neural tract-tracing surface chemistry was chosen for the carrier because the tract-tracers have repeatedly demonstrated efficient reagent uptake and retrograde particle transport, which are believed to be the major barriers to neuronal transfection. Retrograde transport is particularly important for transfection of mature neurons with gene constructs, due to the highly elongated nature of neuronal processes. The proposed carrier also includes surface functional groups for customizable attachment of targeting ligands. The objectives of this research project are to optimize the reagent formulation to achieve a transfection efficiency of greater than 50% in mature mammalian neurons, demonstrate neuronal sub-type targeting capability through ligand attachment, and predictably modify the functional properties of a CNS circuit in an animal model. Successful development of this technology will enable the targeted regulation/observation of neuroplasticity and circuit function in the intact nervous system using molecular constructs. An unprecedented level of transfection efficiency and targeting capability for mature neurons is expected from this nucleic acid carrier by incorporating the essential features of nanoparticulate neural tract-tracers. A combination of high efficiency transfection and precise in vivo targeting capability would serve as a bridge between the field of molecular neurosciences and the systems, behavioral, and preclinical neurosciences, greatly increasing our means to understand and manipulate neuroplasticity and brain function, by enabling the increasingly powerful molecular- biological tools in use today to be broadly applied to neuroscience research. Ultimately, this strategy is expected to lead to more effective therapeutic treatments for a variety of neurological diseases and disorders, including Parkinson's disease, brain injury, and chronic drug addiction.